This project seeks to address several major areas identified in this U01 program. The proposed, highly stable 3-nm ultra-fine iron oxide nanoparticle (uIONP) platform is the next generation of magnetic nanoparticles with the smallest size of its kind for enhanced tumor penetration and retention and the largest per unit surface areas for drug loading. Its novel T1/T2 dual-contrast is switchable depending on its compartmentalization, e.g., dispersed in vessels and self-assembled in tumors, which enables report of the systemic drug delivery process via MRI. The combination of developed uIONPs with the newly developed anti-fouling stealth coating technology and new strategy of targeting both the tumor microenvironment (stroma) and cancer cells will allow us to investigate nanoparticle delivery mechanisms and their implications on therapeutic responses. More importantly targeted theranostic uIONPs carrying anti-cancer agents can overcome the challenge and failure of current therapies in treating pancreatic cancer by breaking the tumor's physiological barriers in stroma and disrupt the tumor microenvironment, leading to improved cancer molecular targeting and targeted drug delivery. The proposed investigation and development of a stroma-breaking theranostic uIONP platform will profoundly change the current nanomaterials' poor efficiency when delivering drugs into fibrotic human cancers, especially pancreatic cancer which is the worst cancer type for drug delivery due to a high content of dense fibrotic tumor stromal and extracellular matrix. With our new understanding of pancreatic cancer biology and expertise in image-guided theranostic IONP delivery systems, in Aim 1 we will develop and characterize dual contrast theranostic uIONPs with stealth antifouling coating (uIONP) that is functionalized with a new uPAR targeting ligand containing the fused peptide of the amino terminal fragment of uPA and the catalytic domain of MMP14 (ATFmmp), and IGF-1 to simultaneously target two cellular receptors, uPAR and IGF- 1R, co-expressed in tumor cells and tumor associated fibroblasts and macrophages. Aim 2 will investigate and determine the stroma breaking ability and efficiency of targeted delivery of ATFmmp-IGF-uIONP in vivo. Developed ATFmmp-IGF-uIONP (alone or with drugs) should be able to extravasate from leaky tumor blood vessels easier and faster than larger IONPs, break extracellular matrix and destroy stromal cells to reach cancer cells, followed by cell receptor-mediated internalization. We will examine the efficiency of targeted delivery and intratumoral distribution of the uIONP in orthotopic human pancreatic cancer patient tissue- derived xenograft (PDX) and transgenic mouse pancreatic tumor models using MR and optical imaging in vivo, multiphoton microscopy ex vivo, and histological and chemical analyses in vitro. In Aim 3, we will determine the improvement in the therapeutic effect and capability of MRI-guided delivery of stroma-breaking theranostic uIONPs carrying cisplatin alone first, and then combined with SN38, which is a camptothecin analogue, as a combination therapy against drug resistance in human pancreatic cancer PDX models.